Issue published February 15, 2023 Previous issue | Next issue
- Volume 133, Issue 4
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On the cover: Ubiquitin-conjugating enzyme E2C in lung cancer
Zhang et al. report that ubiquitin-conjugating enzyme E2C (UBE2C) plays an oncogenic role by promoting the ubiquitylation and degradation of the tumor suppressor DEPTOR by coupling with the APC/CCDH1 E3 ligase. On the cover:
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Review
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Abstract
Kidney disease is a major driver of mortality among patients with diabetes and diabetic kidney disease (DKD) is responsible for close to half of all chronic kidney disease cases. DKD usually develops in a genetically susceptible individual as a result of poor metabolic (glycemic) control. Molecular and genetic studies indicate the key role of podocytes and endothelial cells in driving albuminuria and early kidney disease in diabetes. Proximal tubule changes show a strong association with the glomerular filtration rate. Hyperglycemia represents a key cellular stress in the kidney by altering cellular metabolism in endothelial cells and podocytes and by imposing an excess workload requiring energy and oxygen for proximal tubule cells. Changes in metabolism induce early adaptive cellular hypertrophy and reorganization of the actin cytoskeleton. Later, mitochondrial defects contribute to increased oxidative stress and activation of inflammatory pathways, causing progressive kidney function decline and fibrosis. Blockade of the renin-angiotensin system or the sodium-glucose cotransporter is associated with cellular protection and slowing kidney function decline. Newly identified molecular pathways could provide the basis for the development of much-needed novel therapeutics.
Authors
Samer Mohandes, Tomohito Doke, Hailong Hu, Dhanunjay Mukhi, Poonam Dhillon, Katalin Susztak
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Commentaries
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Abstract
Visceral pain associated with irritable bowel syndrome afflicts 15% of the US population. Although treatments are limited, guanylyl cyclase C (GUCY2C) agonists alleviate pain and constipation. Until now, it was assumed that the activation of GUCY2C and production of cGMP in enterocytes stimulated fluid secretion and reduced visceral sensation. The recent discovery that a subtype of enteroendocrine cells (EECs) known as neuropod cells synapse with submucosal neurons unveiled a pathway for communicating gut signals to the nervous system. In this issue of the JCI, Barton et al. report that GUCY2C is enriched in neuropod cells and is involved with sensory nerve firing. Selective deletion of GUCY2C in mouse models suggests that defective GUCY2C neuropod–cell signaling underlies visceral pain. These studies introduce possibilities for dissociating the secretory and analgesic effects of GUCY2C agonism. Although further work remains, unveiling the role of neuropod cells is a major step in understanding visceral pain.
Authors
Rodger A. Liddle
×Abstract
Cardiac healing following acute myocardial infarction (MI) involves the mobilization and activation of immune cells, including macrophages. In the early phase after MI, macrophages adopt a proinflammatory phenotype, while polarizing toward a reparative one in the late stage. Although metabolic reprogramming has been observed during this transition, the mechanistic links to macrophage differentiation are still poorly understood. In this issue of the JCI, Cai, Zhao and colleagues demonstrate that mitochondrial function in macrophages governed the resolution of inflammation and tissue repair by modulating the phagocytic removal of apoptotic cells (so-called efferocytosis) as well as myofibroblast activation. These findings provide important mechanistic insights into the potential relevance of metabolic modulation of macrophage functions following MI, which might lead to alternative therapeutic strategies for MI.
Authors
David Weissman, Christoph Maack
×Abstract
Multiple myeloma is a hematologic malignancy of plasma cells that manifests with bone marrow tumors causing lytic bone lesions. Autologous stem cell transplantation (ASCT) after high-dose chemotherapy and followed by prolonged maintenance therapy with lenalidomide (LEN) is an effective standard-of-care therapy for multiple myeloma. However, most patients ultimately relapse. Rational combination strategies that address immune dysfunction may prolong the durability of ASCT. In this issue of the JCI, Minnie and colleagues investigated the addition of a checkpoint inhibitor to LEN maintenance therapy after ASCT. They found that the immune checkpoint TIGIT was an optimal target in patient samples. In a syngeneic, immunocompetent multiple myeloma mouse model, blockade of TIGIT synergized with LEN maintenance by inducing immune protection, characterized in part by the expansion of polyfunctional T cells in the bone marrow. The treatment enhanced durable antimyeloma efficacy and has translatable implications.
Authors
Erin W. Meermeier, P. Leif Bergsagel
×Abstract
The CD47/signal regulatory protein α (SIRPα) axis, which functions as an inhibitory phagocytosis checkpoint, also serves as a key mediator in cancer immune evasion. Many cancers, including colorectal cancer (CRC), exploit the expression of CD47 to escape phagocytic clearance and activate the innate immune system. Previous work has indicated that distinct paradigms of posttranslational modifications mediate the regulatory mechanisms of the CD47/SIRPα axis. In this issue of the JCI, Li et al. show that neddylation, a ubiquitin-like modification, inactivates Src homology region 2–containing protein tyrosine phosphatase 2 (SHP2), a downstream target of this pathway. They further show that inhibition of SHP2 sensitizes CRC cells to immunotherapies to which they were previously resistant. Collectively, the results underscore the need for cotargeting SHP2 and immune checkpoints (e.g., programmed death 1 [PD1]) in CRC and possibly other immunotherapy-resistant tumors.
Authors
Kristin Huntoon, Wen Jiang, Betty Y.S. Kim
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Research Letter
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Abstract
Authors
YunZu Michele Wang, Cynthia B. Taggart, John F. Huber, Stella M. Davies, David F. Smith, John B. Hogenesch, Christopher E. Dandoy
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Research Articles
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Abstract
Mutations of G protein–coupled receptors (GPCRs) cause various human diseases, but the mechanistic details are limited. Here, we establish p.E303K in the gene encoding the endothelin receptor type A (ETAR/EDNRA) as a recurrent mutation causing mandibulofacial dysostosis with alopecia (MFDA), with craniofacial changes similar to those caused by p.Y129F. Mouse models carrying either of these missense mutations exhibited a partial maxillary-to-mandibular transformation, which was rescued by deleting the ligand endothelin 3 (ET3/EDN3). Pharmacological experiments confirmed the causative ETAR mutations as gain of function, dependent on ET3. To elucidate how an amino acid substitution far from the ligand binding site can increase ligand affinity, we used molecular dynamics (MD) simulations. E303 is located at the intracellular end of transmembrane domain 6, and its replacement by a lysine increased flexibility of this portion of the helix, thus favoring G protein binding and leading to G protein–mediated enhancement of agonist affinity. The Y129F mutation located under the ligand binding pocket reduced the sodium-water network, thereby affecting the extracellular portion of helices in favor of ET3 binding. These findings provide insight into the pathogenesis of MFDA and into allosteric mechanisms regulating GPCR function, which may provide the basis for drug design targeting GPCRs.
Authors
Yukiko Kurihara, Toru Ekimoto, Christopher T. Gordon, Yasunobu Uchijima, Ryo Sugiyama, Taro Kitazawa, Akiyasu Iwase, Risa Kotani, Rieko Asai, Véronique Pingault, Mitsunori Ikeguchi, Jeanne Amiel, Hiroki Kurihara
×Abstract
Obesity is a risk factor for neurodegenerative disease associated with cognitive dysfunction, including Alzheimer’s disease. Low-grade inflammation is common in obesity, but the mechanism between inflammation and cognitive impairment in obesity is unclear. Accumulative evidence shows that quinolinic acid (QA), a neuroinflammatory neurotoxin, is involved in the pathogenesis of neurodegenerative processes. We investigated the role of QA in obesity-induced cognitive impairment and the beneficial effect of butyrate in counteracting impairments of cognition, neural morphology, and signaling. We show that in human obesity, there was a negative relationship between serum QA levels and cognitive function and decreased cortical gray matter. Diet-induced obese mice had increased QA levels in the cortex associated with cognitive impairment. At single-cell resolution, we confirmed that QA impaired neurons, altered the dendritic spine’s intracellular signal, and reduced brain-derived neurotrophic factor (BDNF) levels. Using Caenorhabditis elegans models, QA induced dopaminergic and glutamatergic neuron lesions. Importantly, the gut microbiota metabolite butyrate was able to counteract those alterations, including cognitive impairment, neuronal spine loss, and BDNF reduction in both in vivo and in vitro studies. Finally, we show that butyrate prevented QA-induced BDNF reductions by epigenetic enhancement of H3K18ac at BDNF promoters. These findings suggest that increased QA is associated with cognitive decline in obesity and that butyrate alleviates neurodegeneration.
Authors
Xing Ge, Mingxuan Zheng, Minmin Hu, Xiaoli Fang, Deqin Geng, Sha Liu, Li Wang, Jun Zhang, Li Guan, Peng Zheng, Yuanyi Xie, Wei Pan, Menglu Zhou, Limian Zhou, Renxian Tang, Kuiyang Zheng, Yinghua Yu, Xu-Feng Huang
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Despite major advances in acute interventions for myocardial infarction (MI), adverse cardiac remodeling and excess fibrosis after MI causing ischemic heart failure (IHF) remain a leading cause of death worldwide. Here we identify a profibrotic coagulation signaling pathway that can be targeted for improved cardiac function following MI with persistent ischemia. Quantitative phosphoproteomics of cardiac tissue revealed an upregulated mitogen-activated protein kinase (MAPK) pathway in human IHF. Intervention in this pathway with trametinib improves myocardial function and prevents fibrotic remodeling in a murine model of non-reperfused MI. MAPK activation in MI requires myeloid cell signaling of protease-activated receptor 2 linked to the cytoplasmic domain of the coagulation initiator tissue factor (TF). They act upstream of pro-oxidant NOX2 NADPH oxidase, ERK1/2 phosphorylation, and activation of profibrotic TGF-β1. Specific targeting with the TF inhibitor nematode anticoagulant protein c2 (NAPc2) starting 1 day after established experimental MI averts IHF. Increased TF cytoplasmic domain phosphorylation in circulating monocytes from patients with subacute MI identifies a potential thromboinflammatory biomarker reflective of increased risk for IHF and suitable for patient selection to receive targeted TF inhibition therapy.
Authors
Venkata Garlapati, Michael Molitor, Thomas Michna, Gregory S. Harms, Stefanie Finger, Rebecca Jung, Jeremy Lagrange, Panagiotis Efentakis, Johannes Wild, Maike Knorr, Susanne Karbach, Sabine Wild, Ksenija Vujacic-Mirski, Thomas Münzel, Andreas Daiber, Moritz Brandt, Tommaso Gori, Hendrik Milting, Stefan Tenzer, Wolfram Ruf, Philip Wenzel
×LepRb+ cell–specific deletion of Slug mitigates obesity and nonalcoholic fatty liver disease in mice
Abstract
Leptin exerts its biological actions by activating the long-form leptin receptor (LepRb). LepRb signaling impairment and leptin resistance are believed to cause obesity. The transcription factor Slug — also known as Snai2 — recruits epigenetic modifiers and regulates gene expression by an epigenetic mechanism; however, its epigenetic action has not been explored in leptin resistance. Here, we uncover a proobesity function of neuronal Slug. Hypothalamic Slug was upregulated in obese mice. LepRb+ cell–specific Slug-knockout (SlugΔLepRb) mice were resistant to diet-induced obesity, type 2 diabetes, and liver steatosis and experienced decreased food intake and increased fat thermogenesis. Leptin stimulated hypothalamic Stat3 phosphorylation and weight loss to a markedly higher level in SlugΔLepRb than in Slugfl/fl mice, even before their body weight divergence. Conversely, hypothalamic LepRb+ neuron–specific overexpression of Slug, mediated by AAV-hSyn-DIO-Slug transduction, induced leptin resistance, obesity, and metabolic disorders in mice on a chow diet. At the genomic level, Slug bound to and repressed the LepRb promoter, thereby inhibiting LepRb transcription. Consistently, Slug deficiency decreased methylation of LepRb promoter H3K27, a repressive epigenetic mark, and increased LepRb mRNA levels in the hypothalamus. Collectively, these results unravel what we believe to be a previously unrecognized hypothalamic neuronal Slug/epigenetic reprogramming/leptin resistance axis that promotes energy imbalance, obesity, and metabolic disease.
Authors
Min-Hyun Kim, Yuan Li, Qiantao Zheng, Lin Jiang, Martin G. Myers Jr., Wen-Shu Wu, Liangyou Rui
×Abstract
Autologous stem cell transplantation (ASCT) with subsequent lenalidomide maintenance is standard consolidation therapy for multiple myeloma, and a subset of patients achieve durable progression-free survival that is suggestive of long-term immune control. Nonetheless, most patients ultimately relapse, suggesting immune escape. TIGIT appears to be a potent inhibitor of myeloma-specific immunity and represents a promising new checkpoint target. Here we demonstrate high expression of TIGIT on activated CD8+ T cells in mobilized peripheral blood stem cell grafts from patients with myeloma. To guide clinical application of TIGIT inhibition, we evaluated identical anti-TIGIT antibodies that do or do not engage FcγR and demonstrated that anti-TIGIT activity is dependent on FcγR binding. We subsequently used CRBN mice to investigate the efficacy of anti-TIGIT in combination with lenalidomide maintenance after transplantation. Notably, the combination of anti-TIGIT with lenalidomide provided synergistic, CD8+ T cell–dependent, antimyeloma efficacy. Analysis of bone marrow (BM) CD8+ T cells demonstrated that combination therapy suppressed T cell exhaustion, enhanced effector function, and expanded central memory subsets. Importantly, these immune phenotypes were specific to the BM tumor microenvironment. Collectively, these data provide a logical rationale for combining TIGIT inhibition with immunomodulatory drugs to prevent myeloma progression after ASCT.
Authors
Simone A. Minnie, Olivia G. Waltner, Kathleen S. Ensbey, Stuart D. Olver, Alika D. Collinge, David P. Sester, Christine R. Schmidt, Samuel R.W. Legg, Shuichiro Takahashi, Nicole S. Nemychenkov, Tomoko Sekiguchi, Gregory Driessens, Ping Zhang, Motoko Koyama, Andrew Spencer, Leona A. Holmberg, Scott N. Furlan, Antiopi Varelias, Geoffrey R. Hill
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The role of tumor-associated macrophages (TAMs), along with the regulatory mechanisms underlying distinct macrophage activation states, remains poorly understood in prostate cancer (PCa). Herein, we report that PCa growth in mice with macrophage-specific Ubc9 deficiency is substantially suppressed compared with that in wild-type littermates, an effect partially ascribed to the augmented CD8+ T cell response. Biochemical and molecular analyses revealed that signal transducer and activator of transcription 4 (STAT4) is a crucial UBC9-mediated SUMOylation target, with lysine residue 350 (K350) as the major modification site. Site-directed mutation of STAT4 (K350R) enhanced its nuclear translocation and stability, thereby facilitating the proinflammatory activation of macrophages. Importantly, administration of the UBC9 inhibitor 2-D08 promoted the antitumor effect of TAMs and increased the expression of PD-1 on CD8+ T cells, supporting a synergistic antitumor efficacy once it combined with the immune checkpoint blockade therapy. Together, our results demonstrate that ablation of UBC9 could reverse the immunosuppressive phenotype of TAMs by promoting STAT4-mediated macrophage activation and macrophage–CD8+ T cell crosstalk, which provides valuable insights to halt the pathogenic process of tumorigenesis.
Authors
Jun Xiao, Fei Sun, Ya-Nan Wang, Bo Liu, Peng Zhou, Fa-Xi Wang, Hai-Feng Zhou, Yue Ge, Tian-Tian Yue, Jia-Hui Luo, Chun-Liang Yang, Shan-Jie Rong, Ze-Zhong Xiong, Sheng Ma, Qi Zhang, Yang Xun, Chun-Guang Yang, Yang Luan, Shao-Gang Wang, Cong-Yi Wang, Zhi-Hua Wang
×Abstract
Innate immune cells play important roles in tissue injury and repair following acute myocardial infarction (MI). Although reprogramming of macrophage metabolism has been observed during inflammation and resolution phases, the mechanistic link to macrophage phenotype is not fully understood. In this study, we found that myeloid-specific deletion (mKO) of mitochondrial complex I protein, encoded by Ndufs4, reproduced the proinflammatory metabolic profile in macrophages and exaggerated the response to LPS. Moreover, mKO mice showed increased mortality, poor scar formation, and worsened cardiac function 30 days after MI. We observed a greater inflammatory response in mKO mice on day 1 followed by increased cell death of infiltrating macrophages and blunted transition to the reparative phase during post-MI days 3–7. Efferocytosis was impaired in mKO macrophages, leading to lower expression of antiinflammatory cytokines and tissue repair factors, which suppressed the proliferation and activation of myofibroblasts in the infarcted area. Mitochondria-targeted ROS scavenging rescued these impairments, improved myofibroblast function in vivo, and reduced post-MI mortality in mKO mice. Together these results reveal a critical role of mitochondria in inflammation resolution and tissue repair via modulation of efferocytosis and crosstalk with fibroblasts. These findings have potential significance for post-MI recovery as well as for other inflammatory conditions.
Authors
Shanshan Cai, Mingyue Zhao, Bo Zhou, Akira Yoshii, Darrian Bugg, Outi Villet, Anita Sahu, Gregory S. Olson, Jennifer Davis, Rong Tian
×Abstract
Pathological neovascularization in age-related macular degeneration (nvAMD) drives the principal cause of blindness in the elderly. While there is a robust genetic association between genes of innate immunity and AMD, genome-to-phenome relationships are low, suggesting a critical contribution of environmental triggers of disease. Possible insight comes from the observation that a past history of infection with pathogens such as Chlamydia pneumoniae, or other systemic inflammation, can predispose to nvAMD in later life. Using a mouse model of nvAMD with prior C. pneumoniae infection, endotoxin exposure, and genetic ablation of distinct immune cell populations, we demonstrated that peripheral infections elicited epigenetic reprogramming that led to a persistent memory state in retinal CX3CR1+ mononuclear phagocytes (MNPs). The immune imprinting persisted long after the initial inflammation had subsided and ultimately exacerbated choroidal neovascularization in a model of nvAMD. Single-cell assay for transposase-accessible chromatin sequencing (scATAC-seq) identified activating transcription factor 3 (ATF3) as a central mediator of retina-resident MNP reprogramming following peripheral inflammation. ATF3 polarized MNPs toward a reparative phenotype biased toward production of proangiogenic factors in response to subsequent injury. Therefore, a past history of bacterial endotoxin–induced inflammation can lead to immunological reprograming within CNS-resident MNPs and aggravate pathological angiogenesis in the aging retina.
Authors
Masayuki Hata, Maki Hata, Elisabeth M.M.A. Andriessen, Rachel Juneau, Frédérique Pilon, Sergio Crespo-Garcia, Roberto Diaz-Marin, Vera Guber, Francois Binet, Frédérik Fournier, Manuel Buscarlet, Caroline Grou, Virginie Calderon, Emilie Heckel, Heather J. Melichar, Jean-Sebastien Joyal, Ariel M. Wilson, Przemyslaw Sapieha
×Abstract
Genetic variants in the third intron of the PRDM6 gene have been associated with BP traits in multiple GWAS. By combining fine mapping, massively parallel reporter assays, and gene editing, we identified super enhancers that drive the expression of PRDM6 and are partly regulated by STAT1 as the causal variants for hypertension. The heterozygous disruption of Prdm6 in mice expressing Cre recombinase under the control of mouse smooth muscle cell protein 22-α promoter (Prdm6fl/+ SM22-Cre) exhibited a markedly higher number of renin-producing cells in the kidneys at E18.5 compared with WT littermates and developed salt-induced systemic hypertension that was completely responsive to the renin inhibitor aliskiren. Strikingly, RNA-Seq analysis of the mouse aortas identified a network of PRDM6-regulated genes that are located in GWAS-associated loci for blood pressure, most notably Sox6, which modulates renin expression in the kidney. Accordingly, the smooth muscle cell–specific disruption of Sox6 in Prdm6fl/+ SM22-Cre mice resulted in a dramatic reduction of renin. Fate mapping and histological studies also showed increased numbers of neural crest–derived cells accompanied by increased collagen deposition in the kidneys of Prdm6fl/+ Wnt1Cre-ZsGreen1Cre mice compared with WT mice. These findings establish the role of PRDM6 as a regulator of renin-producing cell differentiation into smooth muscle cells and as an attractive target for the development of antihypertensive drugs.
Authors
Kushan L. Gunawardhana, Lingjuan Hong, Trojan Rugira, Severin Uebbing, Joanna Kucharczak, Sameet Mehta, Dineth R. Karunamuni, Brenda Cabera-Mendoza, Neeru Gandotra, Curt Scharfe, Renato Polimanti, James P. Noonan, Arya Mani
×Abstract
Our understanding of neuropathic itch is limited due to a lack of relevant animal models. Patients with cutaneous T cell lymphoma (CTCL) experience severe itching. Here, we characterize a mouse model of chronic itch with remarkable lymphoma growth, immune cell accumulation, and persistent pruritus. Intradermal CTCL inoculation produced time-dependent changes in nerve innervations in lymphoma-bearing skin. In the early phase (20 days), CTCL caused hyperinnervations in the epidermis. However, chronic itch was associated with loss of epidermal nerve fibers in the late phases (40 and 60 days). CTCL was also characterized by marked nerve innervations in mouse lymphoma. Blockade of C-fibers reduced pruritus at early and late phases, whereas blockade of A-fibers only suppressed late-phase itch. Intrathecal (i.t.) gabapentin injection reduced late-phase, but not early-phase, pruritus. IL-31 was upregulated in mouse lymphoma, whereas its receptor Il31ra was persistently upregulated in Trpv1-expressing sensory neurons in mice with CTCL. Intratumoral anti–IL-31 treatment effectively suppressed CTCL-induced scratching and alloknesis (mechanical itch). Finally, i.t. administration of a TLR4 antagonist attenuated pruritus in early and late phases and in both sexes. Collectively, we have established a mouse model of neuropathic and cancer itch with relevance to human disease. Our findings also suggest distinct mechanisms underlying acute, chronic, and neuropathic itch.
Authors
Ouyang Chen, Qianru He, Qingjian Han, Kenta Furutani, Yun Gu, Madelynne Olexa, Ru-Rong Ji
×Abstract
Diabetic nephropathy (DN) is a polygenic disorder with few risk variants showing robust replication in large-scale genome-wide association studies. To understand the role of DNA methylation, it is important to have the prevailing genomic view to distinguish key sequence elements that influence gene expression. This is particularly challenging for DN because genome-wide methylation patterns are poorly defined. While methylation is known to alter gene expression, the importance of this causal relationship is obscured by array-based technologies since coverage outside promoter regions is low. To overcome these challenges, we performed methylation sequencing using leukocytes derived from participants of the Finnish Diabetic Nephropathy (FinnDiane) type 1 diabetes (T1D) study (n = 39) that was subsequently replicated in a larger validation cohort (n = 296). Gene body–related regions made up more than 60% of the methylation differences and emphasized the importance of methylation sequencing. We observed differentially methylated genes associated with DN in 3 independent T1D registries originating from Denmark (n = 445), Hong Kong (n = 107), and Thailand (n = 130). Reduced DNA methylation at CTCF and Pol2B sites was tightly connected with DN pathways that include insulin signaling, lipid metabolism, and fibrosis. To define the pathophysiological significance of these population findings, methylation indices were assessed in human renal cells such as podocytes and proximal convoluted tubule cells. The expression of core genes was associated with reduced methylation, elevated CTCF and Pol2B binding, and the activation of insulin-signaling phosphoproteins in hyperglycemic cells. These experimental observations also closely parallel methylation-mediated regulation in human macrophages and vascular endothelial cells.
Authors
Ishant Khurana, Harikrishnan Kaipananickal, Scott Maxwell, Sørine Birkelund, Anna Syreeni, Carol Forsblom, Jun Okabe, Mark Ziemann, Antony Kaspi, Haloom Rafehi, Anne Jørgensen, Keith Al-Hasani, Merlin C. Thomas, Guozhi Jiang, Andrea O.Y. Luk, Heung Man Lee, Yu Huang, Yotsapon Thewjitcharoen, Soontaree Nakasatien, Thep Himathongkam, Christopher Fogarty, Rachel Njeim, Assaad Eid, Tine Willum Hansen, Nete Tofte, Evy C. Ottesen, Ronald C.W. Ma, Juliana C.N. Chan, Mark E. Cooper, Peter Rossing, Per-Henrik Groop, Assam El-Osta
×Abstract
Insulin and IGF-1 receptors (IR and IGF1R) are highly homologous and share similar signaling systems, but each has a unique physiological role, with IR primarily regulating metabolic homeostasis and IGF1R regulating mitogenic control and growth. Here, we show that replacement of a single amino acid at position 973, just distal to the NPEY motif in the intracellular juxtamembrane region, from leucine, which is highly conserved in IRs, to phenylalanine, the highly conserved homologous residue in IGF1Rs, resulted in decreased IRS-1/PI3K/Akt/mTORC1 signaling and increased Shc/Gab1/MAPK cell cycle signaling. As a result, cells expressing L973F-IR exhibited decreased insulin-induced glucose uptake, increased cell growth, and impaired receptor internalization. Mice with knockin of the L973F-IR showed similar alterations in signaling in vivo, and this led to decreased insulin sensitivity, a modest increase in growth, and decreased weight gain when mice were challenged with a high-fat diet. Thus, leucine-973 in the juxtamembrane region of the IR acts as a crucial residue differentiating IR signaling from IGF1R signaling.
Authors
Hirofumi Nagao, Weikang Cai, Bruna B. Brandão, Nicolai J. Wewer Albrechtsen, Martin Steger, Arijeet K. Gattu, Hui Pan, Jonathan M. Dreyfuss, F. Thomas Wunderlich, Matthias Mann, C. Ronald Kahn
×Abstract
Aberrant immune responses to resident microbes promote inflammatory bowel disease and other chronic inflammatory conditions. However, how microbiota-specific immunity is controlled in mucosal tissues remains poorly understood. Here, we found that mice lacking epithelial expression of microbiota-sensitive histone deacetylase 3 (HDAC3) exhibited increased accumulation of commensal-specific CD4+ T cells in the intestine, provoking the hypothesis that epithelial HDAC3 may instruct local microbiota-specific immunity. Consistent with this, microbiota-specific CD4+ T cells and epithelial HDAC3 expression were concurrently induced following early-life microbiota colonization. Further, epithelium-intrinsic ablation of HDAC3 decreased commensal-specific Tregs, increased commensal-specific Th17 cells, and promoted T cell–driven colitis. Mechanistically, HDAC3 was essential for NF-κB–dependent regulation of epithelial MHC class II (MHCII). Epithelium-intrinsic MHCII dampened local accumulation of commensal-specific Th17 cells in adult mice and protected against microbiota-triggered inflammation. Remarkably, HDAC3 enabled the microbiota to induce MHCII expression on epithelial cells and limit the number of commensal-specific T cells in the intestine. Collectively, these data reveal a central role for an epithelial histone deacetylase in directing the dynamic balance of tissue-intrinsic CD4+ T cell subsets that recognize commensal microbes and control inflammation.
Authors
Emily M. Eshleman, Tzu-Yu Shao, Vivienne Woo, Taylor Rice, Laura Engleman, Bailey J. Didriksen, Jordan Whitt, David B. Haslam, Sing Sing Way, Theresa Alenghat
×Abstract
Ubiquitin-conjugating enzyme E2C (UBE2C) mediates ubiquitylation chain formation via the K11 linkage. While previous in vitro studies showed that UBE2C plays a growth-promoting role in cancer cell lines, the underlying mechanism remains elusive. Still unknown is whether and how UBE2C plays a promoting role in vivo. Here we report that UBE2C was indeed essential for growth and survival of lung cancer cells harboring Kras mutations, and UBE2C was required for KrasG12D-induced lung tumorigenesis, since Ube2c deletion significantly inhibited tumor formation and extended the lifespan of mice. Mechanistically, KrasG12D induced expression of UBE2C, which coupled with APC/CCDH1 E3 ligase to promote ubiquitylation and degradation of DEPTOR, leading to activation of mTORC signaling. Importantly, DEPTOR levels fluctuated during cell cycle progression in a manner dependent on UBE2C and CDH1, indicating their physiological connection. Finally, Deptor deletion fully rescued the tumor inhibitory effect of Ube2c deletion in the KrasG12D lung tumor model, indicating a causal role of Deptor. Taken together, our study shows that the UBE2C/CDH1/DEPTOR axis forms an oncogene and tumor suppressor cascade that regulates cell cycle progression and autophagy and validates UBE2C an attractive target for lung cancer associated with Kras mutations.
Authors
Shizhen Zhang, Xiahong You, Yawen Zheng, Yanwen Shen, Xiufang Xiong, Yi Sun
×Abstract
Background Assessing circadian rhythmicity from infrequently sampled data is challenging; however, these types of data are often encountered when measuring circadian transcripts in hospitalized patients.Methods We present ClinCirc. This method combines 2 existing mathematical methods (Lomb-Scargle periodogram and cosinor) sequentially and is designed to measure circadian oscillations from infrequently sampled clinical data. The accuracy of this method was compared against 9 other methods using simulated and frequently sampled biological data. ClinCirc was then evaluated in 13 intensive care unit (ICU) patients as well as in a separate cohort of 29 kidney-transplant recipients. Finally, the consequences of circadian alterations were investigated in a retrospective cohort of 726 kidney-transplant recipients.Results ClinCirc had comparable performance to existing methods for analyzing simulated data or clock transcript expression of healthy volunteers. It had improved accuracy compared with the cosinor method in evaluating circadian parameters in PER2:luc cell lines. In ICU patients, it was the only method investigated to suggest that loss of circadian oscillations in the peripheral oscillator was associated with inflammation, a feature widely reported in animal models. Additionally, ClinCirc was able to detect other circadian alterations, including a phase shift following kidney transplantation that was associated with the administration of glucocorticoids. This phase shift could explain why a significant complication of kidney transplantation (delayed graft dysfunction) oscillates according to the time of day kidney transplantation is performed.Conclusion ClinCirc analysis of the peripheral oscillator reveals important clinical associations in hospitalized patients.Funding UK Research and Innovation (UKRI), National Institute of Health Research (NIHR), Engineering and Physical Sciences Research Council (EPSRC), National Institute on Academic Anaesthesia (NIAA), Asthma+Lung UK, Kidneys for Life.
Authors
Peter S. Cunningham, Gareth B. Kitchen, Callum Jackson, Stavros Papachristos, Thomas Springthorpe, David van Dellen, Julie Gibbs, Timothy W. Felton, Anthony J. Wilson, Jonathan Bannard-Smith, Martin K. Rutter, Thomas House, Paul Dark, Titus Augustine, Ozgur E. Akman, Andrew L. Hazel, John F. Blaikley
×Abstract
SIPRα on macrophages binds with CD47 to resist proengulfment signals, but how the downstream signal of SIPRα controls tumor-infiltrating macrophages (TIMs) is still poorly clarified. Here, we report that the CD47/signal regulatory protein α (SIRPα) axis requires the deneddylation of tyrosine phosphatase SHP2. Mechanistically, Src homology region 2–containing protein tyrosine phosphatase 2 (SHP2) was constitutively neddylated on K358 and K364 sites; thus, its autoinhibited conformation was maintained. In response to CD47-liganded SIRPα, SHP2 was deneddylated by sentrin-specific protease 8 (SENP8), which led to the dephosphorylation of relevant substrates at the phagocytic cup and subsequent inhibition of macrophage phagocytosis. Furthermore, neddylation inactivated myeloid-SHP2 and greatly boosted the efficacy of colorectal cancer (CRC) immunotherapy. Importantly, we observed that supplementation with SHP2 allosteric inhibitors sensitized immune treatment–resistant CRC to immunotherapy. Our results emphasize that the CRC subtype that is unresponsive to immunotherapy relies on SIRPαhiSHP2hiNEDD8lo TIMs and highlight the need to further explore the strategy of SHP2 targeting in CRC therapy.
Authors
Yiqing Li, Hui Zhou, Pan Liu, Dandan Lv, Yichun Shi, Bufu Tang, Jiaqi Xu, Tingting Zhong, Wangting Xu, Jie Zhang, Jianying Zhou, Kejing Ying, Yongchao Zhao, Yi Sun, Zhinong Jiang, Hongqiang Cheng, Xue Zhang, Yuehai Ke
×Abstract
BACKGROUND Mosaic and consensus HIV-1 immunogens provide two distinct approaches to elicit greater breadth of coverage against globally circulating HIV-1 and have shown improved immunologic breadth in nonhuman primate models.METHODS This double-blind randomized trial enrolled 105 healthy HIV-uninfected adults who received 3 doses of either a trivalent global mosaic, a group M consensus (CON-S), or a natural clade B (Nat-B) gp160 env DNA vaccine followed by 2 doses of a heterologous modified vaccinia Ankara–vectored HIV-1 vaccine or placebo. We performed prespecified blinded immunogenicity analyses at day 70 and day 238 after the first immunization. T cell responses to vaccine antigens and 5 heterologous Env variants were fully mapped.RESULTS Env-specific CD4+ T cell responses were induced in 71% of the mosaic vaccine recipients versus 48% of the CON-S recipients and 48% of the natural Env recipients. The mean number of T cell epitopes recognized was 2.5 (95% CI, 1.2–4.2) for mosaic recipients, 1.6 (95% CI, 0.82–2.6) for CON-S recipients, and 1.1 (95% CI, 0.62–1.71) for Nat-B recipients. Mean breadth was significantly greater in the mosaic group than in the Nat-B group using overall (P = 0.014), prime-matched (P = 0.002), heterologous (P = 0.046), and boost-matched (P = 0.009) measures. Overall T cell breadth was largely due to Env-specific CD4+ T cell responses.CONCLUSION Priming with a mosaic antigen significantly increased the number of epitopes recognized by Env-specific T cells and enabled more, albeit still limited, cross-recognition of heterologous variants. Mosaic and consensus immunogens are promising approaches to address global diversity of HIV-1.TRIAL REGISTRATION ClinicalTrials.gov NCT02296541.FUNDING US NIH grants UM1 AI068614, UM1 AI068635, UM1 AI068618, UM1 AI069412, UL1 RR025758, P30 AI064518, UM1 AI100645, and UM1 AI144371, and Bill & Melinda Gates Foundation grant OPP52282.
Authors
Kristen W. Cohen, Andrew Fiore-Gartland, Stephen R. Walsh, Karina Yusim, Nicole Frahm, Marnie L. Elizaga, Janine Maenza, Hyman Scott, Kenneth H. Mayer, Paul A. Goepfert, Srilatha Edupuganti, Giuseppe Pantaleo, Julia Hutter, Daryl E. Morris, Stephen C. De Rosa, Daniel E. Geraghty, Merlin L. Robb, Nelson L. Michael, Will Fischer, Elena E. Giorgi, Harman Malhi, Michael N. Pensiero, Guido Ferrari, Georgia D. Tomaras, David C. Montefiori, Peter B. Gilbert, M. Juliana McElrath, Barton F. Haynes, Bette T. Korber, Lindsey R. Baden, the NIAID HVTN 106 Study Group
×Abstract
Type 2 diabetes (T2D) is caused by insufficient insulin secretion from pancreatic β cells. To identify candidate genes contributing to T2D pathophysiology, we studied human pancreatic islets from approximately 300 individuals. We found 395 differentially expressed genes (DEGs) in islets from individuals with T2D, including, to our knowledge, novel (OPRD1, PAX5, TET1) and previously identified (CHL1, GLRA1, IAPP) candidates. A third of the identified expression changes in islets may predispose to diabetes, as expression of these genes associated with HbA1c in individuals not previously diagnosed with T2D. Most DEGs were expressed in human β cells, based on single-cell RNA-Seq data. Additionally, DEGs displayed alterations in open chromatin and associated with T2D SNPs. Mouse KO strains demonstrated that the identified T2D-associated candidate genes regulate glucose homeostasis and body composition in vivo. Functional validation showed that mimicking T2D-associated changes for OPRD1, PAX5, and SLC2A2 impaired insulin secretion. Impairments in Pax5-overexpressing β cells were due to severe mitochondrial dysfunction. Finally, we discovered PAX5 as a potential transcriptional regulator of many T2D-associated DEGs in human islets. Overall, we have identified molecular alterations in human pancreatic islets that contribute to β cell dysfunction in T2D pathophysiology.
Authors
Karl Bacos, Alexander Perfilyev, Alexandros Karagiannopoulos, Elaine Cowan, Jones K. Ofori, Ludivine Bertonnier-Brouty, Tina Rönn, Andreas Lindqvist, Cheng Luan, Sabrina Ruhrmann, Mtakai Ngara, Åsa Nilsson, Sevda Gheibi, Claire L. Lyons, Jens O. Lagerstedt, Mohammad Barghouth, Jonathan L.S. Esguerra, Petr Volkov, Malin Fex, Hindrik Mulder, Nils Wierup, Ulrika Krus, Isabella Artner, Lena Eliasson, Rashmi B. Prasad, Luis Rodrigo Cataldo, Charlotte Ling
×Abstract
Disorders of isolated mineralocorticoid deficiency, which cause potentially life-threatening salt-wasting crisis early in life, have been associated with gene variants of aldosterone biosynthesis or resistance; however, in some patients no such variants are found. WNT/β-catenin signaling is crucial for differentiation and maintenance of the aldosterone-producing adrenal zona glomerulosa (zG). Herein, we describe a highly consanguineous family with multiple perinatal deaths and infants presenting at birth with failure to thrive, severe salt-wasting crises associated with isolated hypoaldosteronism, nail anomalies, short stature, and deafness. Whole exome sequencing revealed a homozygous splice variant in the R-SPONDIN receptor LGR4 gene (c.618-1G>C) regulating WNT signaling. The resulting transcripts affected protein function and stability and resulted in loss of Wnt/β-catenin signaling in vitro. The impact of LGR4 inactivation was analyzed by adrenal cortex–specific ablation of Lgr4, using Lgr4fl/fl mice mated with Sf1:Cre mice. Inactivation of Lgr4 within the adrenal cortex in the mouse model caused decreased WNT signaling, aberrant zonation with deficient zG, and reduced aldosterone production. Thus, human LGR4 mutations establish a direct link between LGR4 inactivation and decreased canonical WNT signaling, which results in abnormal zG differentiation and endocrine function. Therefore, variants in WNT signaling and its regulators should systematically be considered in familial hyperreninemic hypoaldosteronism.
Authors
Cécily Lucas, Kay-Sara Sauter, Michael Steigert, Delphine Mallet, James Wilmouth, Julie Olabe, Ingrid Plotton, Yves Morel, Daniel Aeberli, Franca Wagner, Hans Clevers, Amit V. Pandey, Pierre Val, Florence Roucher-Boulez, Christa E. Flück
×Abstract
Visceral pain (VP) is a global problem with complex etiologies and limited therapeutic options. Guanylyl cyclase C (GUCY2C), an intestinal receptor producing cyclic GMP(cGMP), which regulates luminal fluid secretion, has emerged as a therapeutic target for VP. Indeed, FDA-approved GUCY2C agonists ameliorate VP in patients with chronic constipation syndromes, although analgesic mechanisms remain obscure. Here, we revealed that intestinal GUCY2C was selectively enriched in neuropod cells, a type of enteroendocrine cell that synapses with submucosal neurons in mice and humans. GUCY2Chi neuropod cells associated with cocultured dorsal root ganglia neurons and induced hyperexcitability, reducing the rheobase and increasing the resulting number of evoked action potentials. Conversely, the GUCY2C agonist linaclotide eliminated neuronal hyperexcitability produced by GUCY2C-sufficient — but not GUCY2C-deficient — neuropod cells, an effect independent of bulk epithelial cells or extracellular cGMP. Genetic elimination of intestinal GUCY2C amplified nociceptive signaling in VP that was comparable with chemically induced VP but refractory to linaclotide. Importantly, eliminating GUCY2C selectively in neuropod cells also increased nociceptive signaling and VP that was refractory to linaclotide. In the context of loss of GUCY2C hormones in patients with VP, these observations suggest a specific role for neuropod GUCY2C signaling in the pathophysiology and treatment of these pain syndromes.
Authors
Joshua R. Barton, Annie K. Londregan, Tyler D. Alexander, Ariana A. Entezari, Shely Bar-Ad, Lan Cheng, Angelo C. Lepore, Adam E. Snook, Manuel Covarrubias, Scott A. Waldman
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ISSN: 0021-9738 (print), 1558-8238 (online)